Previous funding by an R03 application allowed us to develop novel methods that permit the amplification of Plasmodium sequences from fecal DNA. Screening ape samples from over 50 field sites throughout central Africa, we found that P. falciparum is of gorilla origin, and not of chimpanzee, bonobo or ancient human origin. We also found that chimpanzees and gorillas harbor at least nine Plasmodium species, including parasites that are near identical to P. vivax, P. ovale and P. malariae. Given the magnitude of this Plasmodium reservoir and the fact that gorilla P. falciparum has crossed the species to humans already once, the question arises whether additional cross-infections of ape Plasmodium parasites are occurring. This is of critical public health importance, not because such transmissions would be expected to contribute to current malaria morbidity and mortality, but because they would give an indication of the potential of ape malaria parasites to colonize humans should a reduction of P. falciparum transmission rates generate a new ecological niche. In this application, we will examine whether wild apes serve as a recurrent source of human malaria. We will continue to screen wild ape populations for Plasmodium infections, and test humans who live in close proximity to these apes for evidence of cross-species infection. Our hypothesis is that ape parasites have the potential to infect humans but fail to establish persistent infections because of the predominance of P. falciparum. Determining the types, locations and frequencies of ape Plasmodium cross-species infections will be critical to gauge the success of future P. falciparum eradication campaigns. 1. To determine the prevalence of P. vivax and other non-Laverania species in wild apes. We will determine the prevalence, host specificities and distribution of non-Laverania parasites in wild apes, and determine whether chimpanzees and/or gorillas represent a reservoir for human P. vivax in west central Africa. 2. To determine whether wild chimpanzees or gorillas serve as recurrent sources of human infection. We will use ultradeep (454) sequencing to screen humans who live in close proximity to wild apes for evidence of zoonotic Plasmodium infections. This approach will identify Laverania and non-Laverania parasites even if they infect humans at very low frequencies and in the context of multispecies infections. 3. To determine the natural history of ape Plasmodium infections. We will prospectively follow Plasmodium infected sanctuary apes to examine their clinical status, validate our non-invasive detection methods, and obtain blood samples for ape Plasmodium isolation and whole genome sequencing studies. 4. To determine the biological properties that distinguish human P. falciparum from related ape species. We will express ape Plasmodium erythrocyte binding ligands and test their ability to bind to human erythrocytes in order to investigate the molecular basis of Plasmodium host specificity and to assess which ape Plasmodium species have the capacity to cause a blood stage infection in humans.